Impact of minimal inhibitory concentration breakpoint of amoxicillin and clarithromycin on the efficacy of these containing regimens for Helicobacter pylori eradication / 中华消化杂志

Objective:To evaluate the impact of amoxicillin and clarithromycin resistance on the eradication rate of Helicobacter pylori ( H. pylori), and to explore the optimal minimal inhibitory concentration (MIC) breakpoint of amoxicillin and clarithromycin. Methods:From March 2008 to December 2010, patients with H. pylori positive received standard triple therapy to eradicate H. pylori were retrospectively analyzed, 140 patients with H. pylori infetion were included, of which 12 patients did not receive eradication treatment. At 8 to 12 weeks after treatment, the eradication rate of H. pylori of 140 and 128 patients was calculated by intention-to-treat (ITT) and per-protocol population (PP) analysis, respectively. The correlation between amoxicillin and clarithromycin resistance and failure of H. pylori eradication was analyzed. And the relation between different MIC breakpoints of amoxicillin and clarithromycin and failure of H. pylori eradication was also analyzed. Binary logistic regression analysis and consistency test were used for statistical analysis. Results:The results of ITT and PP analysis indicated that the eradication rate of H. pylori of the standard triple therapy was 66.4%(93/140)and 72.7% (93/128), respectively, 95% confidence interval ( CI) 59.3% to 74.3%, and 65.6% to 79.7%, respectively. The results of binary logistic regression analysis showed that amoxicillin resistance (odds ratio ( OR)=6.326, 95% CI 1.090 to 36.725, P=0.040) and clarithromycin resistance ( OR=10.686, 95% CI 4.031 to 28.326, P<0.01) were both independent risk factors of H. pylori eradication failure. The results of consistency test demonstrated that when the MIC breakpoint of amoxicillin was 0.125 mg/L, the correlation between amoxicillin resistance and H. pylori eradication failure was the highest (fair consistency, P<0.05); when the MIC breakpoint of clarithromycin was 2.000 mg/L, the correlation between clarithromycin resistance and H. pylori eradication failure was the highest (moderate consistency, P<0.05). Conclusions:The eradication rate of H. pylori of standard triple therapy dropped to <80%. The decrease of H. pylori eradication rate was related to the resistance of amoxicillin and clarithromycin. The best MIC breakpoints of amoxicillin and clarithromycin were 0.125 and 2.000 mg/L, respectively.

Analysis for common chromosomal breakpoint regions among 586 carriers of reciprocal translocations from Henan Province / 中华医学遗传学杂志

Objective@#To determine the frequency, common chromosomal karyotypes and breakpoints, and involved regions among carriers of reciprocal translocations from Henan Province, and to explore the influence of common breakpoint regions on pregnancy and fetal development.@*Methods@#For 586 carriers of reciprocal translocations, the above features were retrospectively analyzed.@*Results@#The 586 reciprocal translocations were identified among 62 477 subjects, which yielded a frequency of 0.94%. Among these, 572 (0.92%) had abnormal fertility, and 14 (0.02%) had a history of abnormal fetal development. Statistical analysis showed that chromosomes 1, 4, 7 and 11 were most frequently involved, with t(11; 22)(q25; q13) being the most common type of translocation. In total 437 breakpoint regions were identified, with 11q23, 22q13 and 1p36 being most frequently involved, which resulted in infertility, abortion, embryo death, congenital malformation, development delay, mental retardation or a normal phenotype.@*Conclusion@#Above results indicated a 0.92% carrier rate for reciprocal chromosomal translocations in Henan. The location of breakpoint regions may affect the pregnancy and/or fetal development. Discovery of such regions may enable more accurate genetic, reproductive and developmental counseling for carriers, and provide reference for delineation of function and pathogenetic mechanism of the relevant genes.

Análisis de variación del número de copias y de patrones de metilación en la región 15q11-q13 / Variation analysis of the number of copies and methylene patterns in region 15q11-q13

La región q11-q13 del cromosoma 15 humano es proclive a sufrir alteraciones genéticas. Algunos genes de la región presentan expresión parental diferencial monoalélica, regulada por imprinting (EI). Errores en la regulación del EI, disomías uniparentales (DSU), así como también el cambio en el número de copias genómicas (CNV) producidos por sitios susceptibles de quiebre cromosómico (BP), producen alteraciones en esta región. Las enfermedades más frecuentes asociadas son el síndrome de Prader-Willi, el síndrome de Angelman y el síndrome de microduplicación 15q11-q13. En el presente trabajo analizamos la región 15q11-q13 por Methyl specific-multiplex ligation-dependent probe amplification (MS-MLPA) en 181 muestras de ADN derivadas a nuestro servicio de análisis genético molecular. En este trabajo mostramos que, de las 181 muestras, 39 presentaron alteraciones detectables por MS-MLPA. El 61.5% (24/39) de esas alteraciones detectadas fueron deleciones, el 5.1% (2/39) duplicaciones y el 33.3%(13/39) DSU/EI. Los CNV fueron 4 veces más frecuentes que las DSU/EI (OR = 4; IC 95%: 1.56-10.25) consistente con la literatura. Entre los CNV, dos casos atípicos permiten postular posibles sitios BP que no han sido informados en la literatura previamente.

Human chromosome 15q11-q13 region is prone to suffer genetic alterations. Some genes of this region have a differential monoallelic imprinting-regulated expression pattern. Defects in imprinting regulation (IE), uniparental disomy (UPD) or copy number variation (CNV) due to chromosomal breakpoints (BP) in 15q11-q13 region, are associated with several diseases. The most frequent are Prader-Willi syndrome, Angelman syndrome and 15q11-q13 microduplication syndrome. In this work, we analyzed DNA samples from 181 patients with phenotypes which were compatible with the above-mentioned diseases, using Methyl specific-multiplex ligation-dependent probe amplification (MS-MLPA). We show that, of the 181 samples, 39 presented alterations detectable by MS-MLPA. Of those alterations, 61.5% (24/39) were deletions, 5.1% (2/39) duplications and 33.3% (13/39) UPD/IE. The CNV cases were 4 times more frequent than UPD/IE (OR= 4; IC 95%: 1.56-10.25), consistent with the literature. Among the CNVs, two atypical cases allow to postulate new possible BP sites that have not been reported previously in the literature.

Prevalence and Antibiotic Susceptibility of Mycoplasma hominis and Ureaplasma urealyticum in Pregnant Women

Mycoplasma hominis (M. hominis) and Ureaplasma urealyticum (U. urealyticum) are important opportunistic pathogens that cause urogenital infections and complicate pregnancy. The aim of this study was to investigate the prevalence, effects on pregnancy outcomes, and antimicrobial susceptibilities of M. hominis and U. urealyticum. We tested vaginal swabs obtained from 1035 pregnant women for the presence of genital mycoplasmas between June 2009 and May 2014. The laboratory and clinical aspects of genital mycoplasmas infection were reviewed retrospectively, and the identification and antimicrobial susceptibility of genital mycoplasmas were determined using the Mycoplasma IST-2 kit. A total of 571 instances of M. hominis and/or U. urealyticum were detected. Of them, M. hominis was detected in two specimens, whereas U. urealyticum was detected in 472 specimens. The remaining 97 specimens were positive for both M. hominis and U. urealyticum. Preterm deliveries were frequently observed in cases of mixed infection of M. hominis and U. urealyticum, and instances of preterm premature rupture of membrane were often found in cases of U. urealyticum. The rates of non-susceptible isolates to erythromycin, empirical agents for pregnant women, showed increasing trends. In conclusion, the prevalence of M. hominis and/or U. urealyticum infections in pregnant women is high, and the resistance rate of antimicrobial agents tends to increase. Therefore, to maintain a safe pregnancy, it is important to identify the isolates and use appropriate empirical antibiotics immediately.

A comprehensive analysis of breakpoint cluster region-abelson fusion oncogene splice variants in chronic myeloid leukemia and their correlation with disease biology.

BACKGROUND: BCR-ABL fusion oncogene is a hallmark of Chronic Myeloid Leukemia (CML). It results due to translocation between chromosome 22 and chromosome 9 [t (9; 22)(q34; q11)]. It gives rise to translation of a 210 KDa chimeric protein (p210), leading to enhanced tyrosine kinase activity and activation of leukemogenic pathways, ultimately causing onset of CML. In case of CML, the classic fusions are b2a2 or b3a2, fusing exon 13 (b2) or exon 14 (b3) of BCR, respectively, to exon 2 (a2) of ABL. The type of BCR-ABL transcripts are thought to be have different prognosis and hence useful in clinical decision-making. The frequencies of different fusion oncogenes associated with leukemia can vary in different ethnic groups and geographical regions due to interplay of genetic variation in different ethnic populations, diverse environmental factors and living style. Moreover, earlier relevant studies from our region were carried out in small subset of patients. Therefore, objective of this study was to find out frequencies of different BCR-ABL splice variants in larger subset of CML patients. METHODS: A nested reverse transcriptase polymerase chain reaction (RT-PCR) was established to detect BCRABL splice variants in 130 CML patients. Sensitivity of RT-PCR and ability to detect BCR-ABL fusion gene in least possible time was studied. RESULTS: BCR-ABL detection using our optimized RTPCR protocol could be completed in 8 hours, starting from RNA extraction to Gel electrophoresis. Sensitivity of RTPCR assay was of the order of 10−6. Out of 130 Pakistani patients, 83 (63.84%) expressed b3a2 while 47 (36.15%) expressed b2a2 transcript. CONCLUSION: Our RT-PCR was proved to be very quick to detect BCR-ABL fusion oncogene in CML patients within one working day. Because of its sensitivity, it can be used to monitor complete molecular response in CML. BCR-ABL RT-PCR and BCR-ABL splice variants frequency in our study differs from other ethnic groups. It shows that ethnic and geographical differences exist in BCR-ABL splice variant frequency, which may have a profound effect on disease biology as well as implications in prognosis and clinical management of BCR-ABL positive leukemias.

Estrategias para la implementación y reporte de los puntos de corte CLSI vigentes y pruebas fenotípicas confirmatorias para BLEE y carbapenemasas en bacilos Gram negativos en laboratorios clínicos de Colombia / Strategies for implementation and reporting of current CLSI Breakpoints and phenotypic confirmatory tests for ESBLs and carbapenemases in Gram-negative bacilli in Colombian clinical laboratories

En 2010, el Instituto Americano de Estándares Clínicos y de Laboratorio (CLSI) inició un proceso de revisión y actualización de los puntos de corte para microdilución y disco difusión para cefalosporinas (cefazolina, cefotaxima, ceftriaxona, ceftizoxima, ceftazidima), monobactámicos (aztreonam) y carbapenémicos (imipenem, meropenem, ertapenem, doripenem). Los cambios se basaron en modelos PK/PD que buscan predecir la respuesta clínica con el uso exclusivo de la concentración inhibitoria mínima (CIM) y esquemas específicos de dosificación de forma independiente al mecanismo de resistencia expresado. Este nuevo paradigma eliminaría la necesidad de realizar pruebas fenotípicas para beta-lactamasas de espectro extendido (BLEE) y carbapenemasas para tomar decisiones terapéuticas y permitiría utilizarlas únicamente para fines epidemiológicos. Sin embargo, ante las limitaciones de las metodologías actuales para pruebas de susceptibilidad en Colombia, el desconocimiento de estos cambios y la alarma epidemiológica por la aparición de nuevas ß-lactamasas en el país, se hace necesario generar recomendaciones para los laboratorios clínicos, con el fi n de unifi car los criterios para la realización e informe de los antibiogramas en bacilos Gram negativos, incluyendo la implementación de los puntos de corte actuales y la aplicación de las pruebas fenotípicas para la detección de BLEE y carbapenemasas.

In 2010, the Clinical and Laboratory Standards Institute (CLSI) began a process to revise and update the breakpoints for broth microdilution and disk diffusion for cephalosporins (Cefazolin, Cefotaxime, Ceftriaxone, Ceftazidime), monobactams (Aztreonam) and carbapenems (Imipenem, Meropenem, Ertapenem and Doripenem). The changes made were based on PK/PD models that attempt to predict clinical outcomes using minimum inhibitory concentration (MIC) and specific dosage regimens, regardless of the resistance mechanism expressed by the organism. The new breakpoints would eliminate the need to perform screening and confirmatory testing for ESBLs and carbapenemases for treatment decisions, and thus they would be used only for infection control purposes. Nevertheless, there are limitations to current methods in Colombia, a lack of knowledge regarding the recent changes and epidemiologic alarm over new B-lactamases spreading in our country. Therefore it was necessary to formulate and issue recommendations for clinical laboratories, with the aim of standardizing the criteria for reports on antibiograms in Gram-negative bacilli, including the current CLSI breakpoints and applying phenotypic confirmatory testing to detect ESBLs and Carbapenemases.

The inverse PCR:good method for deletion breakpoint mapping / 重庆医学

Objective To map the deletion breakpoint of 9p21 in breast cell line MCF-7 use the inverse PCR .Methods After di-gestion ,ligation and PCR reaction ,the breakpoint was confirmed by sequencing .Results The deletion breakpoint started at chr9 :21819532 and ended at chr9 :21989622 with a small insertion of ACTGG ,which was consistent with the result confirmed by the long range PCR .Conclusion The inverse PCR is one good method for deletion breakpoint mapping and suitable for large sample size detection .

Pitfalls of the Clinical and Laboratory Standards Institute's Revised Breakpoints on Interpretation of the Cephalosporin Susceptibility of an Extended-Spectrum beta-lactamase Producing Klebsiella pneumoniae: Analysis of a 2010 Nationwide Proficiency Surve / 임상검사와정도관리

BACKGROUND: In 2010, the Clinical and Laboratory Standards Institute (CLSI) revised the minimum inhibitory concentration (MIC) breakpoints of cephalosporins and aztreonam to exempt extended-spectrum beta-lactamase (ESBL) confirmatory tests for Enterobacteriaceae. However, the CLSI did not change the MIC breakpoint of cefepime. Here, a proficiency survey of a strain of ESBL-producing Klebsiella pneumoniae was analyzed for MIC distribution and interpretation of cephalosporins and aztreonam. METHODS: The survey strain, K. pneumoniae, which produced SHV-18, was distributed to 170 clinical laboratories as 1 of 5 presumptive clinical specimens through the proficiency survey of the clinical microbiology division of the Korean Association of Quality Assurance for Clinical Laboratories (KAQACL). MIC, zone diameter of inhibition (ZDI), and interpretation of tested antimicrobials, methods of antimicrobial susceptibility testing (AST), and ESBL confirmatory results were collected. RESULTS: According to the revised breakpoints of the 2010 CLSI guidelines, MIC results indicated resistance to aztreonam in 100%, cefepime in 5.5%, cefotaxime in 20%, ceftazidime in 100%, and ceftriaxone in 100% of samples by broth microdilution methods. ZDI results also indicated resistance to aztreonam in 75%, cefepime in 0%, cefotaxime in 66.7%, ceftazidime in 100%, and ceftriaxone in 80% of samples by disk diffusion method. Ninety (75.6%) participants performed an ESBL confirmatory test, and 89 (98.9%) reported ESBL-positive tests. Of the 55 laboratories that tested the susceptibility of cefepime, 50 (90.9%) self-reported to be "resistant" because of ESBL-positive results. CONCLUSIONS: In conclusion, susceptibility testing of ESBL producers against certain cephalosporins is not reliable enough to apply the revised breakpoints presented in the 2010 CLSI guidelines. It is therefore necessary to reach a consensus for interpretation of ASTs of ESBL producers in Korea. Ideally, clinicians should be provided two interpretations based on both the revised breakpoints and ESBL confirmatory testing.

Comparison of Clinical and Laboratory Standards Institute and European Committee on Antimicrobial Susceptibility Testing Breakpoints for beta-Lactams in Enterobacteriaceae Producing Extended-Spectrum beta-Lactamases and/or Plasmid-Mediated AmpC beta-Lacta / 대한임상미생물학회지

BACKGROUND: In 2010, the Clinical and Laboratory Standards Institute (CLSI) revised breakpoints for cephalosporins and carbapenems and indicated that extended-spectrum beta-lactamase (ESBL) testing is no longer necessary for Enterobacteriaceae. We compared the results of the CLSI 2010 and the European Committee on Antimicrobial Susceptibility Testing (EUCAST) MIC breakpoints for Enterobacteriaceae producing ESBL and/or plasmid-mediated AmpC beta-lactamase (PABL). METHODS: A total of 94 well-characterized clinical isolates of Escherichia coli, Klebsiella oxytoca, Klebsiella pneumoniae, Proteus mirabilis, Salmonella spp., Shigella spp., Citrobacter freundii, Enterobacter aerogenes, Enterobacter cloacae, and Serratia marcescens were analyzed. Of them, 57 were ESBL producers, 24 were PABL producers, and 13 were ESBL plus PABL co-producers. Broth microdilution MIC tests were performed for cefotaxime, ceftazidime, aztreonam, cefepime, and imipenem. RESULTS: Among the 94 isolates containing ESBL and/or PABL, the number of isolates that were susceptible to cefotaxime, ceftazidime, aztreonam, cefepime, and imipenem according to the CLSI 2010 vs. the EUCAST breakpoints were 4 (4.3%) vs. 4 (4.3%); 26 (27.7%) vs. 8 (8.5%); 37 (39.4%) vs. 14 (14.9%); 71 (75.5%) vs. 31 (33.0%); and 76 (80.9%) vs. 90 (95.7%), respectively. Of the 18 isolates that were not susceptible to imipenem according to the CLSI 2010 breakpoints, 13 isolates (72.2%) were P. mirabilis. CONCLUSION: The CLSI 2010 MIC breakpoints without tests to detect ESBL and/or PABL for Enterobacteriaceae could be unreliable. Thus, special tests for ESBLs and AmpC beta-lactamases are required to detect the resistance mechanisms involved.

Aminoglycoside Susceptibility Profiles of Enterobacter cloacae Isolates Harboring the aac(6')-Ib Gene / 대한진단검사의학회지

The aminoglycoside 6'-N-acetyltransferases of type Ib (aac(6')-Ib) gene confers resistance to amikacin, tobramycin, kanamycin, and netilmicin but not gentamicin. However, some isolates harboring this gene show reduced susceptibility to amikacin. The European Committee on Antimicrobial Susceptibility Testing (EUCAST) recommends a revision of the phenotypic description for isolates harboring the aac(6')-Ib gene. In this study, we determined the aminoglycoside susceptibility profiles of 58 AAC(6')-Ib-producing Enterobacter cloacae isolates. On the basis of the CLSI and EUCAST breakpoints, a large proportion (84.5% and 55.2%, respectively) of these 58 isolates were found to be susceptible to amikacin. However, among the isolates that were shown to be anikacin-susceptible according to the CLSI and EUCAST breakpoints, only 30.6% and 18.8% isolates, respectively, could be considered to have intermediate resistance on the basis of the EUCAST expert rules. Further studies should be conducted to determine the aminoglycoside susceptibility profiles of aac(6')-Ib-harboring isolates from various geographic regions and to monitor the therapeutic efficacy of amikacin in infections caused by these isolates.

Childhood Chronic Myeloid Leukemia with Monocytosis.

We report two cases of chronic myeloid leukemia (CML) in childhood presenting with monocytosis. History, physical examination and laboratory findings were in favor of juvenile myelomonocytic leukemia in both the cases, but reverse transcriptase polymerase chain reaction (RT-PCR) detected b2a2 and b3a2 transcript of p210 bcr-abl protein characteristic of major BCR breakpoint. Presence of monocytosis in early childhood suggests a viral infection or JMML but a possibility of CML with monocytosis needs to be considered.

Hematological Significance of RT-PCR Test for bcr-abl Rearrangement and the Breakpoint Distribution within the Major bcr in Chronic Myelogenous Leukemia Patients / 대한혈액학회지

BACKGROUND: Bcr-abl rearrangement is the molecular hallmark of chronic myelogenous leukemia (CML). The test for bcr-abl rearrangement, especially using RT-PCR, is the standard test for the diagnosis of CML. We analyzed hematological significances of bcr-abl rearrangement by RT-PCR and the breakpoint distribution within the major bcr in CML patients. METHODS: From 1994 October to 1997 September, we performed the bcr-abl rearrangement using RT-PCR, in 268 untreated patients with various hematologic diseases, and classified the breakpoints within BCR gene as three types (b2a2, b3a2, e1a2) according to PCR product sizes. We compared hematologic parameters between two groups of b2a2 and b3a2 breakpoints in CML. RESULTS: Among the patients with clinically diagnosed CML, 96.8% (61/63) were bcr-abl positive. In ALL, 52.8% (19/36) were bcr- abl positive. All patients with hematologic diseases other than CML or ALL were bcr- abl negative. Among 61 CML patients with positive bcr-abl rearrangement, 19 patients (31.1%) showed b2a2 type and 42 patients (68.9%) b3a2 type. Patients with b3a2 breakpoints showed more frequent peripheral basophilia (P<0.01) than those with b2a2 type. However, other hematologic parameters were not statistically significant. CONCLUSION: RT-PCR test for bcr-abl rearrangement is a specific and efficient test for the diagnosis of CML. However, the hematological significance of b2a2 and b3a2 types is uncertain in CML.

DETECTION OF BCL-2 GENE MAJOR BREAKPOINTREGION REARRANGEMENT IN HUMAN B-CELL LYMPHOMAS / 药物分析学报

Objective To investigate the frequency of t(14;18) in different subtypes of B-cell lymphomas and the ability of the polymerase chain reaction(PCR) to detect this rearrangement in frozen samples. Methods 107 cases of B-cell lymphomas were studied using DNA extracted from fresh-frozen tissues. The DNA samples were amplified by PCR for bcl-2 MBR/JH. The products of bcl-2/JH rearrangement were hybridized with an internal olignucleotide probe of bel-2 MBR. Results The rearranged bel-2MBR/JH gene was detected in 13 of the 25(52. 0％) follicular cen ter lymphomas, according to REAL classification: 8 of 11(72. 7％) grade 1 , 2 of 5(40. 0％) grade Ⅱ , and 3 of 90 (33. 3％) grade Ⅲ. 17 of 82(20. 8％) cases of diffuse large B-cell lympbomas were found to have detectable bcl-2 MBR/JH rearrangement. Conclusion The frequency of bcl-2 MBR/JH rearrangement in diffuse large B-cell lym phomas is significantly lower than those in follicular center lymphomas(χ2=9. 28, P ＜0. 005), suggesting that bcl 2/JH rearrangements occur mainly in follicular center lymphomas. In addition, the result of reconstruction experi ments suggest that amplification of bcl-2 MBR/JH rearrangements by PCR is both sensitive and specific for detection of t (14 ; 18 ) translocation.

Current concern for community-acquired pneumonia / 中国实用内科杂志

Current concerned topics for community-acquired pneumonia(CAP)have been reviewed.There are 3 aspects,including(1)the modified penicillin susceptibility breakpoint of Streptococcus pneumonia and choice of antibiotics treating CAP.(2)Risk factors,which may influence CAP conditions and pathogens were evaluated.(3)Severe CAP and complicating severe sepsis and controversy of glucocorticoid treatment.